Bioscience Discovery, 10(4):214-221, Oct.

- 2019

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ISSN: 2229-3469 (Print); ISSN: 2231-024X (Online)
Research Article
Spectrum and frequency of Chlorophyll mutation in grass pea
(Lathyrus sativus L)
Kulkarni Ganesh. B.

Associate professor in Botany, J.E.S. R.G. Bagdia Arts, S.B. Lakhotia Commerce and
R. Bezonji Science College, Jalna 431203 (M.S.), India
E-mail: dr.kgb98@gmail.com

Article Info Abstract

Received: 25-06-2019,
Revised: 10-08-2019,
Accepted: 26-08-2019
Keywords:
Grass pea (Lathyrus
sativus L.), EMS, SA,
NMU, Gamma rays,
chlorophyll mutation,
leaf aberrations.
The seeds of grass pea (Lathyrus sativus L.) various chemical mutagen concentrations
(EMS 0.05%, 0.10%, 0.15%; SA 0.005%, 0.010%, 0.015% and NMU 0.15%, 0.30%,
0.45%) and physical mutagen doses (Gamma rays- 15KR, 30KR, 45KR) were treated
in laboratory. The Treated and untreated seeds were sown in the field by CRBD
method. The growth of M1 generation observed the characters and number of mutants
in each population thoroughly. The chlorophyll mutation frequency was calculated on
the basis of plant population with explicit chlorophyll mutation and leaf phenotypic
diversity. A noticeable varied range of chlorophyll and morphological mutants were
observed in the M1 generations. Distinct kinds of spectrum of chlorophyll mutants’
viz. xantha, viridis, albino and chlorina were identified in the treated population.
Prominent phenotypic aberrations were also noted viz. reduction in size of leaves,
obtuse and retuse apex, notch at margin, bilobed leaf, lanceolate leaf, wrinkled leaflet
and triforked leaf, etc. The frequency of leaf morphological changes observed in
almost all chemical and physical mutagen in grass pea.

INTRODUCTION cultivate throughout tropical regions of Africa,

Grass pea (Lathyrus sativus L.) is an annual
winter pulse crop in India occupying an area of
about 1.6 million ha (FAO, 2002) and about nearly
200 species and subspecies. L. sativus is the only
member used as a grain legume (Biswas, 2007).
Cultivation was doing for more than 8000 years, a
very less evolutionary progress and improvement of
this crop was made as a pulse crop mainly due to
narrow genetic variability and unavailability of
marker genes with prominent morphological
manifestations, suitable for comprehensive genetic
studies and breeding for high yield of low seed
neurotoxin (ODAP) lines (Smartt, 1984; Waghmare
and Mehra, 2000).
The member of family Fabaceae
(Leguminoceae), is one of the most ancient
originated and cultivated crop among all the crops
in India (Deka and Sarkar, 1990). It grows and
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central and South America, East and West Africa,
Caribbean, Sudan, Egypt, China, Philippines,
Indonesia and Malaysia. In India, it is cultivated in
Tamil Nadu, Andhra Pradesh, Karnataka, Madhya
Pradesh and Maharashtra as a pulses, vegetable and
forage. It had also been proved an excellent source
of nitrogen fixer, to improve soil fertility, also
useful as antioxidants also (Bradley, 1999).
Grass pea (Lathyrus sativus L.) is a crop of
some economic value, in developing countries
including India, Bangladesh, Nepal, and Ethiopia.
The cultivation practices also noticed in China and
countries of Europe, the Middle East, and Northern
Africa. It serves a variety of purposes including
food, feed and fodder owing in part to its nutritive
qualities. In South Asian countries, grass pea is
commonly grown for both grain and fodder
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 Kulkarni Ganesh. B.
purposes. Animal feed from Lathyrus is usually
composed of ground or split grain or flour and is
used primarily to feed lactating cattle or other draft
dal. animals. Human diets include Lathyrus as
grains that are boiled and then either consumed
whole or processed for split In Indian Continent,
Grass pea is a rabbi-season crop adapted to areas
with arid or semiarid regions, is generally sown in
October or November and harvested in February or
March. Sowing date is largely determined by the
time when the monsoon rains end, soil type and soil
moisture profile during October.
Induction of mutation in recent times
yielded some useful morphological variants and the
some author successfully isolated and studied
inheritance patterns of different flower colour, seed
coat colour and stipule mutations in grass pea
(Talukdar et al., 2001a; Talukdar and Biswas,
2005a, b). The main aim of present investigation is
to inquire the effect of different doses of chemical
as well as physical mutagens on chlorophyll
frequency, its spectrum and leaf morphological
changes in Lathyrus.
MATERIALS AND METHODS
The Lathyrus seeds variety of ‘Mahatherara’ was
procured from the Indira Gandhi Krishi
Vishvavidyalaya, Raipur (C.G.) India, used and
treated by various doses of Chemical viz. EMS,
SA, NMU and physical mutagen Gamma rays
[Co60 gamma source (irradiation source capacity to
release 3000 Ci delivery 7200 r/min curie),
irradiated at the Department of Biophysics,
Mutagenic effectiveness (Physical mutagens) = Mf × 100
K rad.
Mutagenic effectiveness (Chemical mutagens) = Mf × 100
c×t
RESULTS AND DISCUSSION
The induction of chlorophyll deficient
sectors observed in all doses of mutagen. Such
sectors were albino (white), xantha (yellow), viridis
(dull green) and chlorina (light green) types. They
were distributed in the leaf lamina. It had been
observed that the various doses of the mutagens
EMS, SA, NMU and Gamma rays had proved
effective inducing the chlorophyll deficient sectors
in Lathyrus seeds (Table 1 and 2). The frequency of
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Government Institute of Science, Aurangabad.
(M.S.) India].
300 (for three replication) healthy and
uniform size of seeds were pre-soaked in distilled
water for 6 hours, for each dose and concentration,
then theses seeds were immerse in the chemical
mutagenic solution for 6 hours, with the different
concentration 0.05, 0.10 and 0.15% for EMS; 0.05,
0.010 and 0.015 % for SA and 0.15, 0.30 and
0.45% for NMU, while for the physical mutagen the
dose applied 15, 30 and 45 KR.
After the each chemical treatment, the
treated seeds were washed thoroughly for 1h in
running tap water to terminate the residual effect of
the mutagenic chemicals. After the completion of
the treatment the treated seeds were sown
immediately in the field along with their respective
controls to rise the M1 generation in a completely
randomized block design (CRBD) with three
replications. All the treatments including the
controls were raised adopting a spacing of 30 cm in
between rows and 15 cm in between plants. The
experiment was conducted during Rabi (winter)
season of 2017. Recommended agronomic practices
were employed for field preparation, sowing and
subsequent management of the Lathyrus crop.
The classification and identification of the
chlorophyll mutants was done based on the
nomenclature adopted by Gustafson (1940). The
various chlorophyll deficient sectors and leaf
abnormalities were noted dose wise separately. The
Mutagenic effectiveness and efficiency were
calculated on the basis of formulae suggested by
Konzak et al. (1965).
chlorophyll deficient sector of plants was noticed
highest by EMS and Gamma rays treatment, as
compared to other mutagens. The frequency of
chlorophyll deficient sectors of Lathyrus plants
ranged from 1.74 to 3.14% by EMS treatments,
from 0.93 to 2.23% by SA treatments, 0.89 to
2.45% by NMU and 1.56 to 3.42% by Gamma ray
treatments. Result shows the maximum chlorophyll
and viable mutation frequency in all the treated
seeds of Lathyrus.
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 Bioscience Discovery, 10(4):214-221, Oct. - 2019

Table1. Effect of mutagens on frequency of plants carrying chlorophyll deficient sectors in M1 generation
of Lathyrus sativus L.

Treatment Dose (%) / (KR) Frequency of +SE

chlorophyll
Deficient Sectors%
Control -- -- --
0.05 1.74 0.62
EMS 0.10 2.45 0.58
0.15 3.14 0.82
0.005 0.93 0.95
SA 0.010 1.75 0.76
0.015 2.23 0.88
0.15 0.89 0.67
NMU 0.30 1.73 0.59
0.45 2.45 1.02
15 1.56 0.89
Gamma rays 30 2.38 0.96
45 3.42 0.73

Figure1. Effect of mutagens on frequency of plants carrying chlorophyll deficient sectors in M 1 generation
of Lathyrus sativus L.

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 Kulkarni Ganesh. B.
Figure 2. Effect of mutagens on total frequency percentage of leaf morphological abnormality observed in
M1 generation of Lathyrus sativus L.
The chlorophyll mutations was also recoded
earlier by various researcher in Lentil (Sharma and
Sharma, 1981), in Chickpea (Kharkwal 1998, Khan
et.al 2005, Bara et al., 2017, in Lathyrus (Prasad
and Das, 1980 and Waghmare and Mehra, 2001), in
Vicia beans (Bhat et. al, 2007), in Cowpea (Girija
and Dhanavel, 2009 and Kumar et al., 2010), in
Black Gram (Thilagavathi and Mullainathan, 2009 ;
Lal et al. 2009), in Mungbean (Kumar et al., 2009),
in green gram (Das and Baisakh, 2010), in Urdbean
(Goyal and Khan, 2010), in Trigonella (Vasu and
Hasan, 2011), in horse gram (Kulkarni and Mogle,
2013), in green gram (Vikram et al., 2014), in
Lablab (Monika and Seethraman, 2017 and
Undirwade and Kulkarni, 2018). Chlorophyll
mutation is shows the directly relation of the
mutagenic effect. The occurrence of chlorophyll
deficient mutant was noticed due to change in gene
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and a set of genes responsible for chlorophyll
mutations. This conclusion is confirmed by the
earlier reports of Kumar et al. (2009) in Mungbean,
Monika and Seetharaman, (2017) in Hyacinth bean.
The seeds of Lathyrus treatment of various
chemical concentration and physical mutagen doses
was revealed the some abnormal shapes and size of
leaves (Fig.1). The leaf variations comprised
bilobed leaf (C), reduction in size of leaf (L), obtuse
apex (A), retuse apex (B), notch at margin (C),
sinuate leaf margin, tri apex leaflet (I), lanceolate
(J) and wrinkled leaflet (L,N). The frequency of
plants showed the leaf morphological changes
observed in various doses of mutagens (Table 1 and
Table 2).
Among the four mutagens, the EMS
especially its 0.15% dose and Gamma rays 45KR
succeeded in inducing the highest morphological
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 Bioscience Discovery, 10(4):214-221, Oct. - 2019

Figure1. Effect of of chemical and physical mutagens Chlorophyll deficient sectors in M1 plants,
morphological abnormality found in Lathyrus sativus L.

Figure Description: Frequency of Chlorophyll mutation: Albino (O), Xantha (D, M, N), Chlorina (A,B,P),
Viridis (G,F).
Frequency of Phenotypic abnormality: Obtuse leaf apex(A), Acute leaf apex (B), Notch in center of leaf(C),
bilobed leaf (C), reduction in size of leaf (L), retuse apex (B), notch at margin(C), sinuate leaf margin, tri
apex leaflet (G, I), lanceolate (J) and wrinkled leaflet (L,O, P), Bilobed leaflet (E,F), Reduction in size of
leaf (B,J), Wrinkled leaflet (L,O,P), Triforked leaf (G), Broad leaf (I,O,P), Opposite leaf with abnormal
Shape (K).

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 Kulkarni Ganesh. B.
frequency (3.14 and 3.42 %) of the leaf
abnormalities, respectively. The lowest frequency
of leaf abnormality changes carrying plants (0.93 %
and 0.89 %) could be noted at 0.05% SA and 0.15
% in NMU, respectively.
In present investigation, leaf morphological changes
may pertinent some abnormalities viz. reduction in
size of leaves, various abnormal leaf shapes and
sizes, observed in the M1 generation mostly noticed
in all treated dosesThe frequency of leaf
morphological changes carrying plants showed
correlation with increasing concentration of
mutagen. The changes in leaf morphology had
reported by some worker earliar, in Pea (Gelin,
1954), Soyabean (Zacharias, 1956), in Safflower
Satpute (1994), in mungbean (Singh et al., 2000 and
Sangsiri et al., 2005), Chickpea (More et al., 2011)
and French bean (Mahamune and Kothekar, 2012),
in horse gram (Kulkarni and Mogle, 2013), in green
gram (Vikram et al., 2014), in Lablab purpureus
(Monika and Seethraman, 2017) in chickpea (Bara
et al., 2017) in Hyacinth bean (Undirwade and
Kulkarni, 2018). The disturbances in phytochromes
causes due to abnormalities in leaves is such as,
chromosomal aberrations, mitotic inhibition,
disrupted auxin synthesis and mineral deficiencies,
disturbance in DNA synthesis, enlargement of
palisade, spongy and mesophyll tissues (Undirwade
and Kulkarni, 2018). The formation of triforked
leaves might be developed by the death of apical
cells at meristematic zone which have specific
influences on the development of leaves and leaf
shape.

Table 2: Frequency (percent) and spectrum of morphological mutants induced by various mutagens in
M1 generation of Lathyrus
Mutant type EMS SA NMU Gama rays frequency
Conc. Freq. Conc. Freq. Conc. Freq. Doses Freq. (%) Total In all
(%) (%) (%)
0.05 0.08 -- -- 0.15 0.06 15KR 0.11 0.25
Plant height
0.62
Tall 0.10 0.12 -- -- 0.30 0.11 30KR 0.14 0.37
0.10 0.06 0.010 0.13 -- -- 15KR 0.07 0.26
Dwarf 0.51
0.15 0.12 -- -- -- -- 30KR 0.13 0.25
Growth 0.10 0.02 0.010 0.15 0.15 0.03 15KR 0.04 0.24
habit 0.39
Bushy -- -- -- -- 0.30 0.15 -- -- 0.15
Leaf 0.10 0.08 0.010 0.13 0.15 0.09 15KR 0.11 0.41
Broad leaf 0.96
0.15 0.11 0.015 0.12 0.30 0.17 30KR 0.15 0.55
Narrow leaf 0.10 0.06 0.010 0.27 0.15 0.02 15KR 0.01 0.36 0.82
0.15 0.09 0.015 0.19 0.30 0.12 30KR 0.06 0.46
Opposite -- -- -- -- -- -- 15KR 0.12 0.12 0.39
leaf 0.15 0.02 0.015 0.08 -- -- 30KR 0.17 0.27
Bilobed leaf 0.10 0.07 0.010 0.1 0.15 0.1 15KR 0.21 0.48 1.13
0.15 0.12 0.015 0.12 0.30 0.17 30KR 0.24 0.65
Elongated 0.10 0.04 0.010 0.19 0.15 0.13 30KR 0.06 0.42 1.02
leaf 0.15 0.17 0.015 0.21 0.30 0.14 45 KR 0.08 0.6
Fused 0.10 0.06 -- -- -- -- -- -- 0.06 0.28
leaflet 0.15 0.14 0.015 0.04 -- -- 30KR 0.04 0.22
wrinkled 0.10 0.19 -- -- 0.15 0.02 15KR 0.02 0.23
0.26
leaflet -- -- -- -- 0.30 0.03 30KR -- 0.03
Triforked 0.10 0.08 0.010 0.07 0.15 0.13 15KR 0.09 0.37 0.87
leaf 0.15 0.1 0.015 0.11 0.30 0.17 30KR 0.12 0.5
Single 0.10 0.09 0.010 0.01 0.15 0.12 30KR 0.12 0.34
flower
0.15 0.12 0.015 0.03 0.30 0.14 45 KR 0.13 0.42
0.76

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 Bioscience Discovery, 10(4):214-221, Oct. - 2019
Conclusion:
The effects of chemical and physical
mutagen in chlorophyll in Lathyrus are applied in
identifying the genetic effect of mutagen. Mutations
in the chlorophyll traits might be reflected in
subsequent generations in the form of different
mutants. The morphological frequency of
chlorophyll deficient sectors induced in the present
investigation included agronomical desirable
characters which may possibly be utilized in future
breeding programme.
Acknowledgement:
Authors are thankful to the Principal and
HOD, Botany Department, JES College, Jalna for
providing research facilities. Author is grateful to
UGC WRO Pune, for giving financial support
through sanction of Minor Research Project.
Authors are extremely grateful to the Department of
Biophysics, Government Institute of Science,
Aurangabad (M.S.) for providing Gamma rays
irradiation facility.
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Kulkarni Ganesh. B., 2019. Spectrum and frequency of Chlorophyll mutation in grass pea (Lathyrus
sativus L).. Bioscience Discovery, 10(4):214-221.
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